Bloom’s Syndrome and PICH Helicases Cooperate with Topoisomerase IIα in Centromere Disjunction before Anaphase

Centromeres are specialized chromosome domains that control chromosome segregation during mitosis, but little is known about the mechanisms underlying the maintenance of their integrity. Centromeric ultrafine anaphase bridges are physiological DNA structures thought to contain unresolved DNA catenations between the centromeres separating during anaphase. BLM and PICH helicases colocalize at these ultrafine anaphase bridges and promote their resolution. As PICH is detectable at centromeres from prometaphase onwards, we hypothesized that BLM might also be located at centromeres and that the two proteins might cooperate to resolve DNA catenations before the onset of anaphase. Using immunofluorescence analyses, we demonstrated the recruitment of BLM to centromeres from G2 phase to mitosis. With a combination of fluorescence in situ hybridization, electron microscopy, RNA interference, chromosome spreads and chromatin immunoprecipitation, we showed that both BLM-deficient and PICH-deficient prometaphase cells displayed changes in centromere structure. These cells also had a higher frequency of centromeric non disjunction in the absence of cohesin, suggesting the persistence of catenations. Both proteins were required for the correct recruitment to the centromere of active topoisomerase IIα, an enzyme specialized in the catenation/decatenation process. These observations reveal the existence of a functional relationship between BLM, PICH and topoisomerase IIα in the centromere decatenation process. They indicate that the higher frequency of centromeric ultrafine anaphase bridges in BLM-deficient cells and in cells treated with topoisomerase IIα inhibitors is probably due not only to unresolved physiological ultrafine anaphase bridges, but also to newly formed ultrafine anaphase bridges. We suggest that BLM and PICH cooperate in rendering centromeric catenates accessible to topoisomerase IIα, thereby facilitating correct centromere disjunction and preventing the formation of supernumerary centromeric ultrafine anaphase bridges

Etude structurale et fonctionnelle des antigenes d'histocompatibilite de lasse I murins

SIGLEAvailable from INIST (FR), Document Supply Service, under shelf-number : T 78406 / INIST-CNRS - Institut de l'Information Scientifique et TechniqueFRFranc

Rôle des modifications post-traductionnelles des histones au cours de la mitose

Les modifications post-traductionnelles des histones jouent un rôle très important au cours du cycle cellulaire et notamment au cours de la mitose. Nous avons montré que les activités histone désacétylases (HDAC) sont impliquées dans le fonctionnement du point de contrôle du fuseau mitotique. Nous avons décrit le rôle d'HDAC3 en mitose, dans la cohésion entre chromatides soeurs et la désacétylation de la lysine 4 de l'histone H3. la phosphorylation de la sérine 7 de CENP-A, un homologue de l'histone H3 uniquement présent au niveau du centromère, est également nécessaire au maintien de la cohésion entre chromatides soeurs. Ces résultats semblent mettre en évidence l'existance d'un "code histone" mitotique lié à la cohésion entre chromatides soeurs.RENNES1-BU Santé (352382103) / SudocSudocFranceF

Expérience de la mise en démarche Qualité de l’unité PESSAC. Dans La démarche Qualité dans la recherche publique et l’enseignement supérieur

Expérience de la mise en démarche Qualité de l’unité PESSAC. Dans La démarche Qualité dans la recherche publique et l’enseignement supérieu

Histone Deacetylase Inhibitors Induce Premature Sister Chromatid Separation and Override the Mitotic Spindle Assembly Checkpoint

International audienceAbstract Histone deacetylase inhibitors (HDACI) are powerful antiproliferative drugs, and are currently undergoing clinical trials as antitumor agents. It would be valuable for both cancer therapy and our knowledge of basic cellular processes to understand the mechanisms by which HDACIs block cell proliferation. Most current models postulate that HDACIs allow the reexpression of tumor suppressor genes silenced in cancer cells. However, other mechanisms, distinct from transcription regulation, may participate in HDACI antiproliferative properties. We report that HDACI treatment induces premature sister chromatid separation in cells in which the mitotic spindle assembly checkpoint (SAC) has already been activated. This effect was transcription-independent. In addition, HDACI-treated mitotic cells displayed SAC inactivation characteristics, including anaphase-promoting complex/cyclosome target degradation, cyclin-dependent kinase 1 inactivation, histone H3 dephosphorylation, and loss of the SAC component MAD2 from the kinetochore. Thus, HDAC inhibition renders the SAC ineffective. Our findings help elucidate the molecular mechanisms of proliferative cell death induced by HDACI treatment and may allow new HDACI-based preclinical and clinical trial protocols to be redesigned so as to target mitosis. [Cancer Res 2007;67(13):6360–7

CDK11p58 kinase activity is required to protect sister chromatid cohesion at centromeres in mitosis

International audienceThe cyclin-dependent kinase CDK11p58 is specifically expressed at G2/M phase. CDK11p58 depletion leads to different cell cycle defects such as mitotic arrest, failure in centriole duplication and centrosome maturation and premature sister chromatid separation. We report that, upon CDK11 depletion, loss of sister chromatid cohesion occurs during mitosis but not during G2 phase. CDK11p58 depletion prevents Bub1 and Shugoshin 1 recruitment but has no effect on the dimethylation of histone H3 lysine 4 at centromeres. We also report that a construct expressing a kinase dead version of CDK11p58 fails to prevent CDK11 depletion-induced sister chromatid separation, showing that CDK11p58 kinase activity is required for protection of sister chromatid cohesion at centromeres during mitosis. Thus, CDK11p58 kinase activity appears to be involved in early events in the establishment of the centromere protection machinery

Histone deacetylase 3 is required for centromeric H3K4 deacetylation and sister chromatid cohesion.

International audienc

The histone methyltransferase NSD3 contributes to cohesin loading during mitotic exit

Abstract During the cell cycle, dynamic post-translational modifications modulate the association of the cohesin complex with chromatin. Phosphorylation / dephosphorylation and acetylation / deacetylation of histones and of cohesin components ensure correct establishment of cohesion during S phase and its proper dissolution during mitosis. In contrast, little is known about the contribution of methylation to the regulation of sister chromatid cohesion. We performed a RNA interference-mediated inactivation screen against 14 histone methyltransferases of the SET domain family that highlighted NSD3 as a factor essential for sister chromatid cohesion in mitosis. We established that NSD3 ensures proper level of the cohesin loader MAU2 and of cohesin itself onto chromatin at mitotic exit. Consistent with its implication in the loading of kollerin and cohesin complexes onto chromatin, we showed that NSD3 associates with chromatin in early anaphase prior to that of MAU2 and RAD21 and dissociates from chromatin upon cell’s entry into prophase. Finally, we demonstrated that of the two NSD3 variant that exist in somatic cells, the long form that carries the methyltransferase activity is the one that acts in cohesion regulation. Taken together, these results describe a novel factor associated with histone methylation in cohesin loading

Calpain 2 is required for sister chromatid cohesion.

International audienceCalpains form a family of Ca(2+)-dependent cysteine proteases involved in diverse cellular processes. However, the specific functions of each calpain isoform remain unknown. Recent reports have shown that calpain 2 (Capn2) is essential for cell viability. We have recently shown that Capn2 is a nuclear protease associated with chromosomes during mitosis in mammalian embryonic cells. We now report that Capn2 depletion impairs mitosis and induces apoptosis in murine cells. Low Capn2 levels induce chromosome alignment defects, the loss of histone H3 threonine 3 phosphorylation at centromeres, and premature sister chromatid separation. Thus, Capn2 may play a role in fundamental mitotic functions, such as the maintenance of sister chromatid cohesion

Aurora A-dependent CENP-A phosphorylation at inner centromeres protects bioriented chromosomes against cohesion fatigue

Sustained spindle tension applied to sister centromeres during mitosis leads to loss of sister chromatid cohesion which is known as cohesion fatigue. Here the authors show that Aurora A-dependent phosphorylation of CENP-A at the inner centromeres protects bioriented chromosomes against cohesion fatigue